Summary PhD thesis Taghi Moazzenzade titled "Current-blockade particle-impact electrochemistry: a single-entity approach for digital (bio)sensing
Early-stage cancer detection is crucial for the successful treatment of patients and reduction of the mortality rate. Liquid biopsy as a non-invasive method enables detection of tumor-derived biomarkers in biofluids such as blood, plasma, and urine. However, detecting cancer markers at an early stage is not straightforward because the concentration of biomarkers is very low. Hence, biosensors with a high sensitivity are required for this aim.
Single-entity electrochemistry (SEE) constitutes a class of highly sensitive systems that employ miniaturized electrodes for the detection of individual analytes. Detecting at the single-entity level is the ultimate mass sensitivity that can be imagined for a sensing system. However, this improved sensitivity can be misleading when these methods are employed for the detection of analytes at ultralow concentrations. Employing a miniaturized transducer decreases the probability of interaction between the target and the electrode surface. Hence, despite the high mass sensitivity, employing micro and nanoscale elements for detecting biomolecules cannot achieve biosensing at ultralow concentrations on a practical time scale. Chapter 2 in this thesis proposes parallelization of the miniaturized transducing elements as a methodology to overcome the mass transport limitation in single-entity sensing.
Among SEE methods, particle-blockade impact electrochemistry studies the collision of non-electroactive particles to a micro or nanoelectrode surface, where individual collisions lead to step-like decreases in the current-time response. Experimental chapters of this thesis aimed to study blockade impact electrochemistry, and employ it as a single-entity method for the digital detection of single-strand DNA oligonucleotides.
In Chapter 3, the effects of low supporting electrolyte concentrations were investigated in blockade impact electrochemistry. In this chapter, examining the signal shape and microparticle trajectories provides insights into a fundamental aspect of electrochemical measurements at low salt regimes; electroosmotic flow (EOF). Finite element analysis and simultaneous optical-electrochemical measurement were employed to evaluate the particle trajectory and signal shape at low salt concentrations. The chapter showed how faradaic reactions at low supporting electrolyte concentrations induce convection in the fluid surrounding a UME due to EOF. The results showed that electroosmotic flow can be the dominant form of transport in measurements at low salt concentrations.
Chapter 4 aimed to improve the signal size uniformity in blockade impact electrochemistry. In this chapter, a microfabricated ring UME was developed to possess a uniform current density on the electrode surface. The ring geometry exhibited a narrower distribution of signal sizes and a higher relative size sensitivity compared to the conventional disk UME.
Chapter 5 aimed to develop a mediator-free single-entity electrochemical (bio)sensor. An intrinsic aspect of amperometric electrochemical methods is the use of a redox mediator for signal generation. In this chapter, the oxygen reduction reaction (ORR) was employed in blockade impact electrochemistry for detecting microparticles at physiological salt conditions. The measurements showed that blockade impact can be performed without employing a synthetic redox mediator. Microparticles can block the oxygen reduction reaction (ORR) on the surface of a UME and generate discrete current signals.
In Chapter 6, a particle-based competitive assay was designed for the detection of the ssDNA oligonucleotide where specifically anchored particles can be dissociated from the surface upon target DNA binding. The motivation for this study was to design an electrochemical sensing method that can detect biomolecules in a digital on/off manner. The results showed that employing microparticles as a label in competitive assays requires strategies for suppressing non-specific adsorption.
As a proof of concept, the competitive assay was used for the detection of ssDNA by blockade impact electrochemistry. However, as discussed in Chapter 2, although single-entity electrochemistry can be optimized for biosensing applications, SEE sensors need to be parallelized to accomplish biosensing at ultralow concentrations. Chapter 7 investigates this idea in detail and introduces the concept of digital electrochemical biosensing in which SEE elements are employed as separately addressable transducing elements. This chapter explains that this parallelized SEE sensor can achieve the ultimate concentration sensitivity because of benefits from both the high mass sensitivity in single elements and overcoming the mass transport problem because of the availability of many sensing elements in parallel.